Mechanisms of cisplatin-induced cell death in malignant mesothelioma cells: Role of inhibitor of apoptosis proteins (IAPs) and caspases

Malignant mesothelioma (MM) is an aggressive and highly chemoresistant tumour. Although cisplatin is used in frontline therapy of this disease treatment remains palliative at best. The biochemical pathways activated by cisplatin and the mechanisms of resistance in mesothelioma cells are poorly understood. Overexpression of inhibitor of apoptosis proteins (IAPs) has been described in clinical mesothelioma tumours and proposed as therapeutic targets. In this study, we examined cisplatin-induced cell death pathways and IAPs in three mesothelioma-derived cell lines. Cisplatin induced cell death in mesothelioma cell lines was characterised by biochemical mechanisms classically associated with apoptosis including: mitochondrial depolarisation, phosphatidylserine translocation and caspase activation. Surprisingly mRNA expression of IAPs in mesothelioma was not upregulated relative to primary mesothelial cells except for survivin which was higher in the most resistant cell line. In contrast, protein expression of both XIAP and survivin was upregulated in all mesothelioma cells, consistent with post-translational regulation. Knockdown of either XIAP or survivin by RNAi did not affect the sensitivity to cisplatin in any of the cell lines. Survivin RNAi did, however, inhibit proliferation in the highest expressing cell line, ONE58. The pan-caspase inhibitor z-VAD and the more selective caspase 3/7 inhibitor z-DEVD had no effect upon the sensitivity of any of the cell lines to cisplatin indicating that caspase-independent pathways predominate. The findings of the present study provide insights into cisplatin-induced mechanisms in mesothelioma cells and show that alternative pathways are operating which may provide new options for targeting this extremely resistant tumour

The Adjuvanticity of an O. volvulus-Derived rOv-ASP-1 Protein in Mice Using Sequential Vaccinations and in Non-Human Primates

Adjuvants potentiate antigen-specific protective immune responses and can be key elements promoting vaccine effectiveness. We previously reported that the Onchocerca volvulus recombinant protein rOv-ASP-1 can induce activation and maturation of naïve human DCs and therefore could be used as an innate adjuvant to promote balanced Th1 and Th2 responses to bystander vaccine antigens in mice. With a few vaccine antigens, it also promoted a Th1-biased response based on pronounced induction of Th1-associated IgG2a and IgG2b antibody responses and the upregulated production of Th1 cytokines, including IL-2, IFN-γ, TNF-α and IL-6. However, because it is a protein, the rOv-ASP-1 adjuvant may also induce anti-self-antibodies. Therefore, it was important to verify that the host responses to self will not affect the adjuvanticity of rOv-ASP-1 when it is used in subsequent vaccinations with the same or different vaccine antigens. In this study, we have established rOv-ASP-1's adjuvanticity in mice during the course of two sequential vaccinations using two vaccine model systems: the receptor-binding domain (RBD) of SARS-CoV spike protein and a commercial influenza virus hemagglutinin (HA) vaccine comprised of three virus strains. Moreover, the adjuvanticity of rOv-ASP-1 was retained with an efficacy similar to that obtained when it was used for a first vaccination, even though a high level of anti-rOv-ASP-1 antibodies was present in the sera of mice before the administration of the second vaccine. To further demonstrate its utility as an adjuvant for human use, we also immunized non-human primates (NHPs) with RBD plus rOv-ASP-1 and showed that rOv-ASP-1 could induce high titres of functional and protective anti-RBD antibody responses in NHPs. Notably, the rOv-ASP-1 adjuvant did not induce high titer antibodies against self in NHPs. Thus, the present study provided a sound scientific foundation for future strategies in the development of this novel protein adjuvant

TNFα kinoid vaccination-induced neutralizing antibodies to TNFα protect mice from autologous TNFα-driven chronic and acute inflammation

The proinflammatory cytokine TNFα is a potent mediator of septic shock and a therapeutic target for chronic inflammatory pathologies including rheumatoid arthritis and Crohn's disease. As an alternative to anti-human TNFα (hTNFα) mAbs and other hTNFα blocker approved drugs, we developed an active anti-hTNFα immunotherapy, based on a vaccine comprised of a keyhole limpet hemocyanin-hTNFα heterocomplex immunogen (hTNFα kinoid) adjuvanted in incomplete Freund's adjuvant. In mice transgenic for hTNFα (TTg mice), hTNFα kinoid vaccination elicited high titers of Abs that neutralized hTNFα bioactivities but did not result in a cellular response to hTNFα. The vaccine was safe and effective in two experimental models. Kinoid-immunized but not control TTg mice resisted hTNFα-driven shock in one model and were prevented from spontaneous arthritis, inflammatory synovitis, and articular destruction in a second model. These data demonstrate an anti-cytokine induction of autoimmune protection against both acute and chronic hTNFα exposure. They show that active vaccination against a human cytokine can be achieved, and that the immune response can be effective and safe